Project Summary Owing mostly to timely reperfusion and better treatment options, the prognosis of acute myocardial infarction (MI) has significantly improved. Nevertheless, MI remains a major cause of death and heart failure (HF) in the US and around the world. Therapeutic intervention is critical in the acute phase of MI, but also increasingly needed in the subsequent chronic phase of adverse cardiac remodeling, which ultimately leads to HF in the growing number of survivors of a heart attack. Here, we propose that two novel and distinct modalities targeting the non-enzymatic signaling function of Na/K-ATPase (NKA) ?1 represent novel therapeutic approaches in each of these two phases of disease progression following an acute cardiac ischemic injury. In the acute phase of MI (Aim 1), we propose that activation of cardioprotective signaling through the cardiac NKA/Src receptor by low concentrations of cardiotonic steroids (CTS) represents a novel adjunct conditioning treatment to mitigate reperfusion injury upon percutaneous coronary intervention. The NKA pathway will be targeted with the FDA-approved CTS digoxin at reperfusion to trigger protection through postconditioning (PostC), and myocardial response to ischemia/reperfusion injury with or without digoxin-induced PostC will be compared ex vivo and in vivo in mice. To confirm that digoxin PostC is mediated through NKA and does not result from an off-target effect, digoxin signaling and protection against ischemia-reperfusion will be compared in genetically engineered mice with cardiac-specific NKA ?1 KO and rescue with a mutant defective in NKA ?1/Src interaction. For the increasing number of patients who will survive MI, we further propose that a deleterious cardiac NKA/Src/reactive oxygen species (ROS) loop plays a role in adverse remodeling and can be targeted to slow or blunt the progression to HF (Aim 2). Based on preliminary evidence, systemic treatment with a peptide designed to block the NKA/Src/ROS loop (pNaktide) two days after an experimental acute MI prevents adverse remodeling and progression to HF in the mouse. Cardiac-specific KO and NKA/Src interaction defective mutant mice will be used to obtain direct evidence of the role of cardiac NKA/Src signaling in the pNaktide effect. Functional, histological, and biochemical studies are proposed to further characterize the protection over time and obtain mechanistic insight. Given the high and increasing prevalence of co-morbidities leading to MI (hypertension, obesity, aging), the stakes are high in developing novel therapies to further improve survival rates and quality of life after MI. The proposed studies shall reveal whether the cardiac NKA/Src pathway is a novel, robust and translatable signaling module to target in both the acute phase and chronic phase of MI.